Numerous diseases are caused by gene defects. The goal of gene therapy is the effective treatment or cure of these diseases by the introduction of the normal gene or repair of the gene defect. A large research effort has been mounted to optimize gene transfer vectors mostly based on adenoviruses and retroviruses. Yet, the initial promise of gene therapy has been undermined by the biology of the commonly used viral vectors: retroviruses are intrinsically mutagenic and oncogenic as they integrate into the human genome;vectors derived from adenoviruses induce vigorous humoral and cellular immune responses that negate the therapeutic success. Isogenis is taking a two pronged approach to the problem of immunogenicity in Adenoviral vectors. First, others have also shown that Adenoviral vectors deleted of viral genes, i.e. fully deleted (fd, gutless) are less immunogenic than vectors containing more viral genes. Despite intensive research, the efficient production of fd Adenoviral vectors still requires the use of 'helper'viruses to deliver crucial Adenoviral genes. Isogenis had learned from its discussion with the FDA (Dr. Stephanie Simek, Deputy Director, Division of Cellular and Gene Therapy) that the use of 'helper'viruses for the production of fd Adenoviral vectors was considered problematic as present purification systems did not deliver preparations with 'helper'virus contamination levels of less than 1-in-3 x 1010 particles. Isogenis has produced a novel fd Adenoviral vector production system that avoids the use of a 'helper'virus and replication-efficient recombinants. Isogenis will use this novel vector architecture to produce clinical gene therapy vectors for the treatment of hemophilia A and ornithine transcarbamylase deficiency (OTCD). The activity of these vectors will be tested in pre-clinical animal models. Second, to overcome any remaining immunogenicity, Isogenis has developed innovative immune inhibitory gene transfer vectors. These employ the highly specific, yet effective immune suppression of the natural "veto" immune inhibitory phenomenon. Briefly, cells expressing CD8 on their surface exert a "veto" on T-cells responding to other antigens on their surface. The activity of these vectors will be tested in pre-clinical animal models. The development of these new Adenoviral vectors is a major step forward in getting gene therapy back on track. PUBLIC HEALTH RELEVANCE: The goal of gene therapy is the cure of the disease by the introduction or repair of the defective gene. In the case of hemophilia A and OTCD, the 'missing'enzyme will be directly delivered to the resident hepatocytes. Gene therapy vectors will have to lead to sustained gene expression in their target tissue to provide a permanent cure. Immune responses directed against vector proteins generally negate any therapeutic effect. Therefore, strategies are being sought that allowed the production of effective gene therapy vectors that did no longer contain vector-derived, i.e. viral, genes. In the case of adenoviral vectors such fully deleted vectors had been designed. There efficient production depended on the use of helper viruses. Yet, the use of a helper virus was considered problematic by the FDA. Present purification systems do not deliver preparations with helper virus contamination levels of less than 1-in-3x1010. In this application, we therefore propose to investigate how w novel set of helper-virus independent fully deleted Adenoviral gene transfer vectors function in vivo.